低氧诱导因子1α过表达对PC12细胞在CoCl2拟缺氧模型中凋亡的影响  

Effects of over-expression of hypoxia inducible factor 1α on apoptosis of PC12 cells under hypoxic injury condition mimicked by CoCl2

作　　者：赵宁辉[1] 杨勇涛[1] 徐蔚[1] 郝金刚[2] 赵娜[3] 黄晓玮[3] 

机构地区：[1]昆明医学院第二附属医院神经外科,昆明650101 [2]昆明医学院第二附属医院放射科,昆明650101 [3]云南大学生物资源保护与利用重点实验室,昆明650091

出　　处：《中华神经医学杂志》2010年第12期1189-1192,共4页Chinese Journal of Neuromedicine

基　　金：国家自然科学基金面上项目（30660190）;云南省科技厅-昆明医学院联合专项基金（2008CD021,20080CD19）;云南省中青年学术技术带头人后备人才项目（2009C1034）

摘　　要：目的 研究CoCl2处理拟缺氧损伤条件下低氧诱导因子1α（HIF-1α）过表达对PC12神经细胞凋亡状态的影响.方法 分化后的PC12细胞分为2组,实验组转染pEGFPC1-HIF-1α△ODD表达质粒,对照组转染pEGFPC1.CoCl2处理模拟细胞缺氧损伤条件;Western blotting检测2组细胞常氧和缺氧时HIF-1α的表达;Hochest33342染色、流式细胞仪和Caspase-3活性检测3种方法观察神经细胞拟缺氧损伤时HIF-1α的过表达对细胞凋亡的影响.结果 50 μmol/L及100μmol/L CoCl2处理PC12细胞12 h、24 h可以诱导细胞拟缺氧损伤模型,实验组细胞常氧和缺氧时HIF-1α表达均高于对照组.Hochest33342染色结果提示实验组凋亡细胞明显少于对照组细胞.50 μmol/L CoCl2处理24 h后,流式细胞仪结果显示实验组细胞凋亡率为（5.41±3.29）%,对照组为（8.35±2.59）%,差异有统计学意义（P＜0.05）.加入50μmol/LCoCl2处理12 h后,实验组细胞中Caspase-3活性的增加倍数明显低于实验组细胞.结论 适量CoCl2处理的神经细胞系拟缺氧损伤模型中,HIF-1α的过表达对细胞凋亡有显著的抑制作用.Objective To investigate the effects of over-expression ofhypoxia inducible factor 1α （HIF-1α） on apoptosis of PC12 cells under hypoxic injury condition mimicked by CoCl2. Methods To obtain the neural cell line over-expressing HIF-1α, the recombinant eukaryotic expression vector of pEGFPC1-HIF-1α△ODD was transfected into the differentiated PC12 cells, and pEGFPC1 was transfected into the PC12 cells as control group. PC12 cells under simulative hypoxic injury condition was constructed by CoCl2 treatment. The expression of HIF-1α was analyzed under normal and hypoxia conditions, respectively, using the method of Western blotting. Then Hoechst33342 staining, flow cytometer and caspase-3 activity detection were employed to measure and compare the cell apoptosis rates between the over-expressing HIF-1α cell line and the controls. Results We successfully constructed the model with simulative hypoxic injury to the PC12 cell line via the treatment of 50 μ mol/L or 100 μ mol/L CoCl2 for 12-24 h. The results from Western blotting illustrated that cells transfected by pEGFPC1-HIF-1α△ODD highly expressed HIF-1α under both normal and hypoxic conditions as compared with the control cells. Furthermore, Hoechst33342 staining indicated that the apoptosis rate in those transfected by pEGFPC1-HIF-1α△ODD was significantly decreased as compared with that in the control cells; the apoptosis rate in those transfected by pEGFPC1-HIF-1α△ODD （5.41±3.29）% and the control cells （8.35±2.59）% was obviously different 24 h after treatment with 50 μ mol/L CoCl2 （P〈0.05）.The caspase-3 activities were significantly different between those transfected by pEGFPC1-HIF-1α△ODD （2.152±0.384） and control cells （0.921 ±0.287） 12 h after treatment with 50μmol/L CoCl2 （P〈0.05）. Conclusion In the model of neural cells undergoing simulative hypoxic injury by the treatment of CoCl2, over-expression of HIF-1α can inhibit the cell apoptosis.

关 键 词：细胞低氧 低氧诱导因子1Α 二氯化钴 细胞凋亡 

分 类 号：R686[医药卫生—骨科学]